Submitted on June 28, 2005
Accepted on September 23, 2005
Role of calmodulin methionine residues in mediating the productive association with cardiac ryanodine receptors
Edward M Balog1*, Laura E Norton1, David D Thomas1, and Bradley R Fruen1
1 Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, MN, USA
* To whom correspondence should be addressed. E-mail: ed.balog{at}ap.gatech.edu.
Calmodulin (CaM) binds to the cardiac ryanodine receptor Ca2+ release channel (RyR2) with high affinity, and may act as a regulatory channel subunit. Here we determine the role of CaM Met residues in the productive association of CaM with RyR2, as assessed via determinations of [3H]ryanodine and [35S]CaM binding to cardiac muscle sarcoplasmic reticulum (CSR) vesicles. Oxidation of all nine CaM Met residues abolished the productive association of CaM with RyR2. Substitution of CaM's C-terminal Mets with Leu decreased the extent of CaM inhibition of CSR vesicle[3H]ryanodine binding. In contrast, replacing CaM's N-terminal Mets with Leu increased the concentration of CaM required to inhibit CSR vesicle[3H]ryanodine binding but did not alter the extent of inhibition. Site-specific substitution of individual CaM Met residues with Gln demonstrated that Met 124 was required for both high-affinity CaM binding to RyR2 and for maximal CaM inhibition. These results thus identify a Met residue critical for the high-affinity association of CaM with RyR2 channels.
